It is proposed to continue work on the characterization of isolated acetylcholine receptors using equilibrium binding and kinetic methods. Techniques to be used include nuclear magnetic resonance spectroscopy, electron-spin resonance methods, stop-flow and temperature jump techniques. Protein chemical characterization of isolated receptor molecules will also be conducted. Physical methods will be used to detect and hopefully quantitate conformational changes which may occur upon ligand binding. The relationship of specific ion binding sites to cholinergic ligand sites will also be investigated. A second line of approach for the overall elucidation of the mechanisms of cholinergic synaptic depolarization is to study membrane fragments particularly enriched in cholinergic receptors (i.e., where approximately 50% of the total protein in membrane vesicles corresponds to acetylcholine receptor). Techniques to be used include electron microscopy, x-ray crystallography, nuclear magnetic resonance and electron spin resonance techniques. A third approach consists of reassociation of isolated acetylcholine receptors with phospholipid molecules. The properties of such systems will be investigated by equilibrium binding methods, electron microscopy, and ion flux measurements in reconstituted vesicles.